Several lines of evidence indicate that a differential development of opiate receptor subtypes occurs in neonatal rat brain. We propose to exploit this system to address the question of whether a subcellular population of opiate bindng sites localized in brain smooth microsomes and Golgi represent newly-synthesized receptors in transit from the cell body to terminals. First we will complete our analysis of the sequence of appearance of opiate receptor subtypes in the total membrane populations from forebrain during the postnatal period. Then we will repeat the study using synaptic membranes and microsomes. To accomplish this, procedures developed to isolate these fractions in mature brain will be modified for the developing postnatal central nervous system. The same criteria will be used, i.e., marker enzymes, electron microscopy, bouyant density and binding characteristics, to ensure the relative homogeneity of the two fractions and the absence of cross-contamination. Opiate receptor binding affinities and capacities in microsomal and synaptic membrane preparations from neonatal rats of various ages will be assessed by displacement analysis using combinations of radioactive and unlabeled prototypic ligands for opiate receptor subtypes. We will analyze binding data with a non-linear curve-fittng computer program for multiple site bindings. Techniques of electron microscopic autoradiography and histochemistry will be adopted for fine structual localization of opiate receptors in insolated fractions containing synaptic membranes, Golgi or endoplasmic reticulum. The postnatal appearance of subtypes of opiate receptors in subcellular populations of different brain regions will be correlated with the time course of manifestation of receptor-mediated physiological processes such as analgesia, electrophysiological responses to opiates and opiate regulation of catecholamine turnover.